1. Inflammation
Inflammation is commonly divided into three phases: acute inflammation, the immune response and chronic inflammation. Acute inflammation is the initial response to tissue injury and is mediated by the release of histamine, serotonin, bradykinin, prostaglandins and leukotrienes. The immune response, usually preceded by the acute inflammation phase, occurs when immunologically competent cells are activated in response to foreign organisms or antigenic substances liberated during the acute or chronic inflammatory response. The outcome of the immune response for the host may be beneficial, as it causes invading organisms to be phagocytosed or neutralized. However, the outcome may be deleterious if it leads to chronic inflammation without resolution of the underlying injurious process as occurs in rheumatoid arthritis.
The treatment of patients with inflammation leads to the slowing or arrest of the tissue-damaging process as well as the relief of pain, which is the presenting symptom and the major continuing complaint of the patient.
Anti-inflammatory agents are usually classified as steroidal or glucocorticoids and nonsteroidal anti-inflammatory agents (NSAIDs). The glucocorticoids are powerful anti-inflammatory agents but the high toxicity associated with chronic corticosteroid therapy inhibits their use except in certain acute inflammatory conditions. Therefore, the nonsteroidal anti-inflammatory drugs have assumed a major role in the treatment of chronic conditions such as rheumatoid arthritis.
Among the non-steroidal anti-inflammatory agents are included derivatives of aminoarylcarboxylic acids, arylacetic acids, arylbutyric acids, arylcarboxylic acids, arylpropionic acids, pyrazole, pyrazolone, salicylic acid and some other derivatives of different chemical structure, including specific anti-arthritic/anti-rheumatic agents.
It would be highly desirable to provide new nonsteroidal anti-inflammatory agents that could serve as alternatives to current anti-inflammatory drugs.
2. Vaccines and Adjuvants
Lymphocytes are the central cells of the immune system, responsible for acquired immunity and the immunologic attributes of diversity, specificity, memory, and self/non-self recognition. Mature B cells are distinguished from other lymphocytes by their synthesis and display of membrane-bound immunoglobulin (antibody) molecules, which serve as receptors for antigens. Interaction between antigen and the membrane-bound antibody on a mature naive B cell, results in the activation and differentiation of B-cell clones of corresponding specificity and the consequent production of B cell clones lacking the membrane-bound antibody, but which secrete antibody molecules with the same antigen-binding specificity.
T lymphocytes, like B lymphocytes, have membrane receptors for antigens. However, unlike the membrane-bound antibody on B cells, the T-cell receptor (TCR) does not recognize free antigen. Instead, the TCR recognizes only antigen that is bound to a self-molecule encoded by genes within the major histocompatibility complex (MHC). To be recognized by most T cells, the antigen must be displayed together with MHC molecules on the surface of antigen-presenting cells (APC) or on virus-infected cells, cancer cells, and grafts.
Like B cells, T cells express distinctive membrane molecules. All T-cell subpopulations express the TCR, a complex of polypeptides that includes CD3, and most can be distinguished by the presence of one or the other of two membrane molecules, CD4 and CD8. T cells that express the membrane glycoprotein molecule CD4 are restricted to recognizing antigen bound to class II MHC molecules, whereas T cells expressing CD8, a dimeric membrane glycoprotein, are restricted to recognition of antigen bound to class I MHC molecules.
In general, expression of CD4 and of CD8 also defines two major subpopulations of T lymphocytes. CD4+ T cells generally function as T helper (TH) cells and are class-II restricted; CD8+ T cells generally function as T cytotoxic (TC) cells and are class-I restricted.
TH cells are activated by recognition of an antigen-class II MHC complex on an antigen-presenting cell. After activation, the TH cell begins to divide and gives rise to a clone of effector cells, each specific for the same antigen-class II MHC complex. These TH cells secrete various cytokines, which play a central role in the activation of B cells, T cells, and other cells that participate in the immune response.
Changes in the pattern of cytokines produced by TH cells can change the type of immune response that develops among other leukocytes. Thus TH cells have been divided into two groups by the characteristic cytokines they secrete when activated: the TH1 response produces a cytokine profile that supports inflammation and activates mainly certain T cells and macrophages whereas the TH2 response activates mainly B cells and immune responses that depend upon antibodies. Thus, TH1 cells secrete IL-2, which induces T-cell proliferation, and cytokines such as IFN-γ, which mediates tissue inflammation. TH2 cells, in contrast, secrete IL-4, which activates B cells to secrete antibodies of certain IgG isotypes and suppresses the production of TH1 inflammatory cytokines, and IL-10, which suppresses inflammatory cytokine production by macrophages, and thus indirectly reduces cytokine production by TH1 cells, and affects antigen-presenting cells by down-regulating class II MHC expression.
Autoimmunity results from an inappropriate response of the immune system against self-components leading to activation of self-reactive clones of T or B cells, and generation of humoral or cell-mediated responses against endogenous antigens, with consequent injury to cells, tissues and organs. Sometimes, the damage is caused by antibodies as in the autoimmune disorders Addison's disease, autoimmune anemia, e.g. autoimmune hemolytic anemia and pernicious anemia, Hashimoto's thyroiditis and scleroderma.
Many autoimmune disorders e.g. insulin-dependent diabetes mellitus (IDDM or type I diabetes), multiple sclerosis, rheumatoid arthritis and autoimmune thyroiditis are characterized by tissue destruction mediated by T cells activated by an endogenous antigen. These immune responses to self-antigens are maintained by the persistent activation of the self-reactive T lymphocytes.
Autoimmune diseases can be divided into organ-specific autoimmune diseases, in which the immune response is directed to a target antigen unique to a single organ or gland, so that the manifestations are largely limited to that organ, and systemic autoimmune diseases, in which the response is directed toward a broad range of target antigens and involves a number of organs and tissues. Examples of organ-specific autoimmune diseases include insulin-dependent diabetes mellitus, multiple sclerosis, rheumatoid arthritis, thyroiditis, and myasthenia gravis, and examples of systemic autoimmune diseases include systemic lupus erythematosus and scleroderma.
It is the TH1 cells which contribute to the pathogenesis of organ-specific autoimmune diseases. For example, there is strong evidence that, in mice, experimental autoimmune encephalomyelitis (EAE) is caused by CD4+ TH1 cells specific for the immunizing antigen, e.g. myelin basic protein (MBP) or proteolipid protein (PLP). The disease can be transferred from one animal into another by T cells from animals immunized with either MBP or PLP or by cloned T-cell lines from such animals. TH1-type responses also appear to be involved in other T-cell mediated diseases or conditions such as contact dermatitis.
Most cases of organ-specific autoimmune diseases develop as a consequence of self-reactive CD4+ T cells. Analysis of these T cells revealed that the TH1/TH2 balance can affect whether autoimmunity develops. TH1 cells have been involved in the development of autoimmunity, whereas, in several cases, TH2 cells not only protected against the induction of the disease but also against progression of established disease and in the induction and maintenance of allograft tolerance.
Several therapeutic approaches have been explored for treatment of autoimmune diseases. Identification and sequencing of various autoantigens have led to the development of new approaches to modulate autoimmune T-cell activity. Whole antigens involved in the pathogenesis of the autoimmune disease or peptides derived from their sequences have been proposed for the treatment of autoimmune diseases.
Synthetic peptides suitable for immunologically specific therapy of an autoimmune disease are peptides that are recognized by T cells involved in the pathogenesis of the autoimmune disease. These peptides may have a sequence consisting of a pathogenic sequence within the sequence of an antigen involved in the disease or may be an analogue thereof, in which sequence one or more native amino acid residues are substituted by different amino acid residues, particularly a so-called “altered peptide”, which contains a single amino acid substitution in the epitope of the pathogenic native counterpart (i.e., the region that contacts the TCR), but have no alterations in the agretope (i.e., the region that contacts the MHC).
Each autoimmune disease will have its ideal peptide for use in therapy that is derived directly from the sequence of an antigen associated with the disease, or is an altered peptide, or another analogue thereof. Thus, a disease like multiple sclerosis (MS) involving T cells reactive to self-antigens such as myelin basic protein (MBP), myelin oligodendrocyte glycoprotein (MOG) and proteolipid protein (PLP), will require for its therapy a peptide of MBP, MOG or PLP or an analogue thereof; myasthenia gravis can be treated with a peptide from the acetylcholine receptor; thyroiditis with a peptide from thyroglobulin; diabetes type 1 with a peptide of glutamic acid decarboxylase (GAD) or a peptide from the insulin sequence; systemic lupus erythematosus with a peptide derived from the protein P53; and Guillain-Barré syndrome with a peptide from the myelin antigen P2.
In recent years, peptides derived from a pathogenic self-antigen associated with an autoimmune disease or analogues thereof have been proposed for treatment of the disease. For example, peptides derived from the human MBP sequence (U.S. Pat. No. 5,817,629; U.S. Pat. No. 6,252,040) and analogues thereof (U.S. Pat. No. 5,948,764; U.S. Pat. No. 6,329,499) have been described for treatment of multiple sclerosis; peptide analogues of the 65 kD isoform of human GAD and of insulin have been proposed for treatment of diabetes (U.S. Pat. No. 5,945,401 and U.S. Pat. No. 6,197,926, respectively); and an autoantigen or a fragment thereof have been described for the treatment of uveoretinitis (U.S. Pat. No. 5,961,977). Each and all of the patents cited above are hereby incorporated by reference herein as if fully disclosed herein.
For each of the various autoimmune diseases, it would be desirable to administer the relevant peptide in an adjuvant that would activate T cells of the anti-inflammatory TH2 phenotype. This would be expected to arrest the autoimmune process. There are also situations not involving therapy of an autoimmune disease in which it would be useful to activate specific T cells with a TH2 phenotype. However, treatment involving self-antigens must be done in adjuvants that do not induce TH1-type immunity that might activate dangerous TH1 autoimmunity in the treated subject. Thus, there is a need to identify adjuvants capable of being combined with specific antigens that will induce non-inflammatory TH2-type T cells.
Adjuvants, by their nature, are non-specific immunomodulators. An adjuvant suitable for the purposes outlined above would be a non-specific immunomodulator that could be combined in a therapeutic vaccination with an antigen or other molecule so as to induce the activation of specific T cells of the desired anti-inflammatory phenotype.
Several peptides suitable for the therapy of T-cell mediated diseases, disorders or conditions such as autoimmune diseases were shown to be effective when administered to mice subcutaneously (SC) in an oil vehicle such as an emulsion of mineral oil known as incomplete Freund's adjuvant (IFA). However, IFA as well as complete Freund's adjuvant (CFA; a preparation of mineral oil containing various amounts of killed organisms of Mycobacterium) are not allowed for human use because the mineral oil cannot be degraded in the body.
It would be highly desirable to discover effective vehicles for peptide therapy that would be degradable and act as an adjuvant that serves as a carrier, or depot or immune potentiator/enhancer.
3. References Related to the Art
Some fatty alcohols and esters of fatty acids have been described as solvents or emulsifiers for use in pharmaceutical compositions. For example, cetyl alcohol may be used in pharmaceutical compositions as emulsifying and stiffening agent (The Merck Index, 2001, 13th edition, pp. 347-8, #2037), oleyl alcohol may be used as a carrier for medicaments (The Merck Index, 2001, 13th edition, p. 1222, #6900), and alkyl esters of oleic acid may be used as solvents for medicaments (The Merck Index, 2001, 13th edition, p. 6899, #6898).
A mixture of higher aliphatic primary alcohols, primarily isolated from beeswax, was described as having moderate anti-inflammatory activity. The composition of such a mixture was not disclosed (Rodriguez et al., 1998).
The mass spectra of nicotinates of long chain alcohols, e.g. octadecyl and (Z)-9-octadecen-1-yl nicotinates, have been studied to elucidate the structure of long chain alcohols (Vetter and Meister, 1981). No biologic activity was assigned to the compounds.
Esters of 4-aminomethyl-benzoic acid (PAMBA) with C6-C16 saturated alcohols, e.g. decyl, undecyl, tetradecyl and hexadecyl alcohols, have been tested for their antifibrinolytic activity and found to be not active (Markwardt et al., 1966). PAMBA esters with short chain alcohols were found to be able to decrease the proliferation of in vitro cultivated endothelial cells, the hexyl ester being the more effective (Beyer and Pilgrim, 1991).
Alkyl N,N-disubstituted amino acids, e.g. alkyl N,N-dimethylamino acetate wherein the alkyl is octyl, decyl, dodecyl, or tetradecyl, and decyl(4-methyl-1-piperazinyl)acetate, have been described as transdermal penetration enhancers for indomethacin and possibly for other drugs (Wong et al., 1989; U.S. Pat. No. 4,980,378).
Complexes for use in gene therapy comprising a therapeutically active substance and a cationic lipid such as quaternary piperazinium compounds substituted at both the 1 and 4 positions by a methyl and an oleyloxycarbonylmethyl radicals are described in U.S. Pat. No. 6,291,423.
Esters of N,N-dimethyl-aminoacetic acid with long chain alkanols, e.g. tetradecyl, cetyl and stearyl alcohols, and alkenols, are described in JP 2000-302650 for use in hair cosmetics. The oleyl ester is not specifically disclosed.
Betaine [(carboxymethyl)trimethylammonium hydroxide inner salt] esters with long-chain alcohols such as decyl, lauryl, myristyl, pamityl, stearyl and oleyl alcohol were prepared and their pharmacodynamic properties have been studied (Metayer and Jacob, 1952), or their activity as biocides for cooling water treatment was tested (Rucka et al., 1983).
Quaternary ammonium salts of lauryl, myristyl and cetyl esters of N-carboxymethyl-piperidine, -piperazine and -morpholine compounds were described as germicides (Smith et al., 1951).
Stearyl esters of amino acids, e.g. glycine, phenylglycine, alanine, valine, leucine, lysine, proline, phenylalanine, and tyrosine, and stearyl esters of peptides have been proposed as adjuvants for bacterial and viral human vaccines (Penney et al., 1985, 1993; Nixon-George et al., 1990).
Esters of DL-ω-phenyl-amino acids with C4-C10 alkanols, such as DL-2-phenylglycine octyl or decyl ester or DL-2-(4-dimethylaminophenyl)glycine octyl ester have been described as antiphlogistic, antihistaminic, spasmolytic, antioxidant and anti-inflammatory (Schulz et al., 1982; Schewe et al., 1991; Kontogiorgis et al., 2001).
Higher alkyl esters of amino acid, e.g. lauryl, myristyl, cetyl and stearyl esters of glycine, phenylglycine, alanine, valine, norvaline, leucine, isoleucine, lysine, and phenylalanine, and their N-lower alkyl derivatives are described in U.S. Pat. No. 3,821,403 (Misato et al., 1974) as useful for control of plant diseases.